bims-indpro Biomed News
on Intrinsically disordered proteins
Issue of 2022–01–30
eight papers selected by
Sara Mingu, Johannes Gutenberg University



  1. J Am Chem Soc. 2022 Jan 23.
      Protein folding and dynamics are governed by an intricate interplay of thermal and viscosity-mediated effects. The solvent viscosity contributes to the frictional drag in protein dynamics. In addition to this viscosity-dependent effect, there is also an intriguing viscosity-independent component that represents the intrinsic resistance of the polypeptide chain to changing its conformation. This solvent-independent component is termed internal friction. A longstanding question is what is the fundamental molecular origin of internal friction in highly solvated and rapidly fluctuating intrinsically disordered proteins (IDPs) devoid of any persistent intrachain interactions? Here, we present a unique case to directly demonstrate that sequence-specific backbone dihedral barriers control local internal friction in an archetypal IDP, namely, α-synuclein. We performed site-directed fluorescence depolarization kinetics using picosecond time-resolved fluorescence anisotropy measurements to directly observe the directional decorrelation arising due to short-range backbone torsional fluctuations in the dihedral space. A linear viscosity-dependent model of the dihedral relaxation time yielded a finite zero-viscosity intercept that corresponds to internal friction. Our site-specific dynamic readouts were able to detect localized sequence-specific frictional components that are otherwise skewed in viscosity-dependent long-range chain fluctuations. Our results revealed the presence of low internal friction in nonproline sequence segments. In contrast, a proline introduces torsional stiffness in the segment exhibiting high internal friction that can be compensated by a conformationally flexible glycine. Such an intriguing interplay of local dihedral dynamics can modulate sequence-dependent internal friction in a wide range of IDPs involved in a myriad of important events including folding, binding, assembly, and phase transitions.
    DOI:  https://doi.org/10.1021/jacs.1c11236
  2. Biophys J. 2022 Jan 21. pii: S0006-3495(22)00040-6. [Epub ahead of print]
      Proteins and water couple dynamically over a wide range of time scales. Motivated by their central role in protein function, protein-water dynamics and thermodynamics have been extensively studied for structured proteins, where correspondence to structural features has been made. However, properties controlling intrinsically disordered protein (IDP)-water dynamics are not yet known. We report results of megahertz-to-terahertz dielectric spectroscopy and molecular dynamics simulations of a group of IDPs with varying charge content along with structured proteins of similar size. Hydration water around IDPs is found to exhibit more heterogeneous rotational and translational dynamics compared to water around structured proteins of similar size, yielding on average more restricted dynamics around individual residues of IDPs, charged or neutral, compared to structured proteins. The on-average slower water dynamics is found to arise from excess tightly bound water in the first hydration layer, which is related to greater exposure to charged groups. The more tightly bound water to IDPs correlates with the smaller hydration shell found experimentally, and impacts entropy associated with protein-water interactions, the contribution of which we estimate based on the dielectric measurements and simulations. Water-IDP dynamic coupling at terahertz frequencies is characterized by the dielectric measurements and simulations.
    DOI:  https://doi.org/10.1016/j.bpj.2022.01.011
  3. J Biochem. 2022 Jan 26. pii: mvab156. [Epub ahead of print]
      Stress response is important for sensing and adapting to environmental changes. Recently, RNA-protein condensates, which are a type of membrane-less organelle formed by liquid-liquid phase separation, have been proposed to regulate the stress response. Because RNA-protein condensates are formed through interactions between positively charged proteins and negatively charged RNAs, the ratio of proteins to RNAs is critical for phase-separated condensate formation. In particular, long noncoding RNAs (lncRNAs) can efficiently nucleate phase-separated RNA-protein condensates because of their secondary structure and long length. Therefore, increased attention has been paid to lncRNAs because of their potential role as a regulator of biological condensates by phase separation under stress response. In this review, we summarize the current research on the involvement of lncRNAs in the formation of RNA-protein condensates under stress response. We also demonstrate that lncRNA-driven phase separation provides a useful basis to understanding the response to several kinds of cellular stresses.
    Keywords:  Long noncoding RNA; RNA-binding protein; RNA-protein condensate; phase separation; stress response
    DOI:  https://doi.org/10.1093/jb/mvab156
  4. J Biol Chem. 2022 Jan 22. pii: S0021-9258(22)00073-4. [Epub ahead of print] 101633
      Most transcription factors possess at least one long intrinsically disordered transactivation domain that binds to a variety of co-activators and co-repressors and plays a key role in modulating the transcriptional activity. Despite the crucial importance of these domains, the structural and functional basis of transactivation remains poorly understood. Here, we focused on ATF4/CREB-2, an essential transcription factor for cellular stress adaptation. Bioinformatic sequence analysis of the ATF4 transactivation domain sequence revealed that the first 125 amino acids have noticeably less propensity for structural disorder than the rest of the domain. Using solution NMR spectroscopy complemented by a range of biophysical methods, we found that the isolated transactivation domain is predominantly yet not fully disordered in solution. We also observed that a short motif at the N-terminus of the transactivation domain has a high helical propensity. Importantly, we found that the N-terminal region of the transactivation domain is involved in transient long-range interactions with the basic-leucine zipper domain involved in DNA binding. Finally, in vitro phosphorylation assays with the casein kinase 2 (CK2) show that the presence of the basic-leucine zipper domain is required for phosphorylation of the transactivation domain. This study uncovers the intricate coupling existing between the transactivation and basic-leucine zipper domains of ATF4, highlighting its potential regulatory significance.
    Keywords:  intrinsically disordered protein; protein kinase; solution NMR spectroscopy; transcription factor
    DOI:  https://doi.org/10.1016/j.jbc.2022.101633
  5. J Am Chem Soc. 2022 Jan 24.
      Several point mutations can modulate protein structure and dynamics, leading to different natures. Especially in the case of amyloidogenic proteins closely related to neurodegenerative diseases, structural changes originating from point mutations can affect fibrillation kinetics. Herein, we rationally designed mutant candidates to inhibit the fibrillation process of amyloid-β with its point mutants through multistep in silico analyses. Our results showed that the designed mutants induced kinetic self-assembly suppression and reduced the toxicity of the aggregate. A multidisciplinary biophysical approach with small-angle X-ray scattering, ion mobility-mass spectrometry, mass spectrometry, and additional in silico experiments was performed to reveal the structural basis associated with the inhibition of fibril formation. The structure-based design of the mutants with suppressed self-assembly performed in this study could provide a different perspective for modulating amyloid aggregation based on the structural understanding of the intrinsically disordered proteins.
    DOI:  https://doi.org/10.1021/jacs.1c10173
  6. Front Mol Biosci. 2021 ;8 794646
      Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disorder characterized by the degeneration of both upper and lower motor neurons in the brain and spinal cord. ALS is associated with protein misfolding and inclusion formation involving RNA-binding proteins, including TAR DNA-binding protein (TDP-43) and fused in sarcoma (FUS). The 125-kDa Matrin3 is a highly conserved nuclear DNA/RNA-binding protein that is implicated in many cellular processes, including binding and stabilizing mRNA, regulating mRNA nuclear export, modulating alternative splicing, and managing chromosomal distribution. Mutations in MATR3, the gene encoding Matrin3, have been identified as causal in familial ALS (fALS). Matrin3 lacks a prion-like domain that characterizes many other ALS-associated RNA-binding proteins, including TDP-43 and FUS, however, our bioinformatics analyses and preliminary studies document that Matrin3 contains long intrinsically disordered regions that may facilitate promiscuous interactions with many proteins and may contribute to its misfolding. In addition, these disordered regions in Matrin3 undergo numerous post-translational modifications, including phosphorylation, ubiquitination and acetylation that modulate the function and misfolding of the protein. Here we discuss the disordered nature of Matrin3 and review the factors that may promote its misfolding and aggregation, two elements that might explain its role in ALS pathogenesis.
    Keywords:  ALS; Matrin3; intrinsically disordered domains; protein misfolding; proteinopathy
    DOI:  https://doi.org/10.3389/fmolb.2021.794646
  7. Proc Natl Acad Sci U S A. 2022 Jan 25. pii: e2115523119. [Epub ahead of print]119(4):
      The exceptional elastic resilience of some protein materials underlies essential biomechanical functions with broad interest in biomedical fields. However, molecular design of elastic resilience is restricted to amino acid sequences of a handful of naturally occurring resilient proteins such as resilin and elastin. Here, we exploit non-resilin/elastin sequences that adopt kinetically stabilized, random coil-dominated conformations to achieve near-perfect resilience comparable with that of resilin and elastin. We also show a direct correlation between resilience and Raman-characterized protein conformations. Furthermore, we demonstrate that metastable conformation of proteins enables the construction of mechanically graded protein materials that exhibit spatially controlled conformations and resilience. These results offer insights into molecular mechanisms of protein elastomers and outline a general conformation-driven strategy for developing resilient and functional protein materials.
    Keywords:  conformation; elasticity; polymorphism; protein; silk
    DOI:  https://doi.org/10.1073/pnas.2115523119
  8. Methods Mol Biol. 2022 ;2406 359-370
      Recombinant protein expression in E. coli often induces the expressed protein to accumulate in insoluble aggregates, named inclusion bodies (IBs), that represent easy to isolate, highly pure protein reservoirs. IBs can be solubilized by denaturing agents but this procedure requires, for complex globular proteins, a refolding step that can be challenging. However, the lack of cooperatively folded tertiary structure in intrinsically disordered proteins (IDP) makes them ideal candidates for this purification strategy. Given the wide abundance of IDPs, their relevance in many disease areas and the numerous IDP-associated biological functions, the interest in this class of proteins has increased substantially over the last decade. Here we present a broad and versatile method for the production and isolation of IDPs from inclusion bodies under denaturant conditions that overcomes the challenges associated with the propensity of these sequences to precipitate from solution and becoming proteolytically degraded.
    Keywords:  Aggregation; Amyloid; Chemical denaturation; Denaturing conditions; Inclusion bodies; Intrinsically disordered proteins; Liquid–liquid phase separation; Proteolysis; Recombinant purification; Urea
    DOI:  https://doi.org/10.1007/978-1-0716-1859-2_21